RANK Signaling in Osteoclast Differentiation & Function

破骨细胞分化中的 RANK 信号传导

• 批准号: 6533019
• 负责人: XU FENG
• 金额: $ 28.7万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-09-28 至 2006-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6533019
• 关键词: binding proteins; biological signal transduction; cell differentiation; colony stimulating factor; cytokine receptors; gene targeting; genetically modified animals; laboratory mouse; macrophage; nuclear factor kappa beta; osteoclasts; physiologic bone resorption; protein structure function; tumor necrosis factor alpha

DESCRIPTION (provided by applicant): Osteoclasts (OCs), the principal bone-resorbing cells, arise from hematopoietic precursors of monocyte-macrophage lineage upon stimulation of two key factors: M-CSF and RANKL (also known as OPGL/ODF/TRANCE). RANKL, identified as a member of TNF family, is a potent inducer of OC differentiation, activation and survival. RANKL exerts its effects by binding to its receptor RANK, identified as a member of TNF receptor superfamily, on OC precursors or mature OCs. Both RANKL-/- and RANK-/- mice develop osteopetrosis due to the failure to form OCs, indicating that RANK signaling is essential for differentiation. Furthermore, RANKL activates the resorptive function of the mature OCs and inhibits their apoptosis. Thus, RANKL is pivotal to generation, activation and survival of OCs. However, the specific motifs (residues) in the RANK cytoplasmic domain mediating OC differentiation, activation and survival have not been identified. We have developed a tool which permits us to identify the specific motifs mediating these three effects. Using a retroviral technology, we expressed a chimeric receptor containing the extracellular domain of TNFR1 linked to the transmembrane and cytoplasmic domains of RANK in authentic OC precursors isolated from TNFR1/R2 double knock-out mice. Treatment of the chimera-expressing OC precursors with TNF-alpha as a RANKL surrogate, plus M-CSF, generates OCs, indistinguishable from those induced by RANKL and M-CSF. Thus, using this chimeric protein approach, we are positioned to delineate the specific motifs in the RANK cytoplasmic domain mediating OC differentiation, activation and survival. Since members of TNF receptor superfamily transmit the intracellular signaling by binding intracellular adaptor proteins such as TRAFs, we hypothesize that RANK, as a member of the TNF receptor superfamily, initiates its signaling in OC differentiation, activation and survival by recruiting adaptor proteins, including TRAFs, to specific motifs in the RANK cytoplasmic domain. Our Specific Aims are to identify: (1) specific motifs in the RANK cytoplasmic domain mediating OC differentiation; (2) specific motifs in the RANK cytoplasmic domain mediating OC activation; and (3) specific motifs in the RANK cytoplasmic domain mediating OC survival.

描述（由申请人提供）：破骨细胞（OC），主要成分 骨吸收细胞，由造血前体细胞产生 单核细胞-巨噬细胞谱系在两个关键因子的刺激下：M-CSF和 RANKL（也称为 OPGL/ODF/TRANCE）。 RANKL，被认定为TNF成员 家族，是 OC 分化、激活和存活的有效诱导剂。 RANKL 通过与其受体 RANK 结合来发挥作用，RANK 被确定为 TNF 受体超家族的成员，位于 OC 前体或成熟 OC 上。两个都 RANKL-/- 和 RANK-/- 小鼠由于无法形成 OC 而出现骨石症， 表明 RANK 信号传导对于分化至关重要。此外， RANKL 激活成熟 OC 的再吸收功能并抑制其 细胞凋亡。因此，RANKL 对于细胞的产生、激活和生存至关重要。 OC。然而，RANK 胞质结构域中的特定基序（残基） 介导 OC 分化、激活和存活的机制尚未确定。 我们开发了一种工具，可以让我们识别特定的图案 调节这三种效应。使用逆转录病毒技术，我们表达了 含有 TNFR1 胞外结构域的嵌合受体，该受体与 真实 OC 前体中 RANK 的跨膜和细胞质结构域 分离自 TNFR1/R2 双敲除小鼠。治疗 表达嵌合体的 OC 前体，以 TNF-α 作为 RANKL 替代物，加上 M-CSF 产生 OC，与 RANKL 和 M-CSF 诱导的 OC 没有区别。 因此，利用这种嵌合蛋白方法，我们能够描绘出 RANK细胞质结构域中的特定基序介导OC分化， 激活和生存。由于 TNF 受体超家族的成员传递 通过结合细胞内衔接蛋白（例如 TRAFs，我们假设 RANK 作为 TNF 受体超家族的一员， 通过以下方式启动其在 OC 分化、激活和存活中的信号传导： 将接头蛋白（包括 TRAF）招募到 RANK 中的特定基序 细胞质结构域。我们的具体目标是确定：（1） 介导 OC 分化的 RANK 细胞质结构域； (2)特定主题 在介导 OC 激活的 RANK 细胞质结构域中； (3)特定主题 在介导 OC 存活的 RANK 细胞质结构域中。